Oscillating sensor and fluid sample analysis using an oscillating sensor

ABSTRACT

An assembly design for an oscillating resonator-based sensor where an oscillating crystal resonator such as a quartz crystal resonator is rigidly affixed or ‘mounted’ onto a solid substrate in such a fashion that the resonator can either rest flush against the substrate surface or upon a rigid mounting adhesive. Once cured, the mounting adhesive forms a liquid tight seal between the mounted resonator and the substrate such that only the sensing electrode surface will be exposed to fluids applied to the front side of the substrate. The mounted resonator assembly is designed in such a way that it can be interfaced with a fluid delivery system to form a liquid tight chamber or flow cell around the mounted resonator without incurring additional physical impact upon the mounted resonator. The assembled flow cell can in turn be used to direct multiple fluid streams to flow in a laminar manner over the sensing surface of the mounted resonator and by varying the rates of flow for the different laminar flowing fluid streams the total hydraulic pressure exerted on the surface of the mounted resonator can be held constant.

FIELD OF THE INVENTION

The invention regards a new sensor assembly and method of fluid deliveryto the sensor, and more specifically pertains to a sensor assembly forfluid sample testing that includes a rigid attachment of the coresensing device to a mounting substrate to minimize and control alldirect physical contact to the core sensing device of the assembly inconjunction with a unique method of test fluid delivery to controlpressure fluctuations upon the core sensing device during operation.

BACKGROUND OF INVENTION

Oscillating crystal resonators can be used as very sensitive masssensors in gas and liquid phase. It was shown by Sauerbrey (Sauerbrey,G., Z. Phys. 155 (1959), p. 206-222) that material deposited onto aresonator surface will change the resonators fundamental oscillationfrequency proportional to the mass of the deposited material. Due to theextreme sensitivity of these resonators to changes in mass on theirsurfaces, oscillating crystal resonators can be employed to determinemass changes on a molecular level, and are often referred to as QuartzCrystal Microbalances or QCM. An oscillating crystal resonator generallyconsists of a thin plate of piezoelectric material, such as a quartzcrystal wafer, with metal electrodes deposited on each face of theplate. Applying an electric field between the electrodes, or across thepiezoelectric plate, causes a physical displacement in the piezoelectricmaterial. Due to this “piezoelectric phenomenon” caused the by theelectric current, steady oscillations of piezoelectric plates can beachieved through the application of a stable electric field in thismanner. Once stable, changes in oscillation of the piezoelectric platesdue to the addition or subtraction of mass from their surface can bequantified with great accuracy.

In 1980 researchers (Konash, P. L. and Bastiaans, G. J., Anal Chem. 52(1980), p. 1929-1931), successfully utilized an oscillating crystalresonator as a sensor for measurement in the liquid phase. Over theyears the use of oscillating crystal resonators as acoustic sensors inliquid phase applications have became very popular. Today there arethousands of literature publications documenting the application ofoscillating quartz crystal resonators for use as liquid phase acousticsensors. These sensors have been used to measure the presence and amountof chemical substances such as agricultural pesticides, toxins, and foodadditives in samples. A form of quartz sensor called a biosensor hasbeen used to measure the presence and interactions of proteins, such asantibodies and hormones, nucleic acids, as well as pharmaceutical drugs,in liquid samples ranging from bodily fluids to organic solvents.

When operating an oscillating crystal resonators as a liquid phasesensor it is a requirement that the liquid sample interact with only oneof the electrode coated surfaces on the resonator. The reasons for thisare two-fold: 1) to eliminate electrical short-circuits between theelectrodes of the resonator, and 2) to minimize loss of the resonatorQ-factor (ratio of stored energy to dissipated energy in thepiezoelectric plate) due to the liquids viscosity. To overcome theproblem of creating a short-circuit between the electrodes the resonatormount design must isolate the back (driving/non-sensing) electrode fromthe front (sensing) electrode such that only the sensing electrode isexposed to the liquid under test. The viscosity of liquids cansignificantly dampen or even completely stop the oscillation of acrystal resonator. The higher the viscous load on the resonator thelower its sensitivity to changes in mass on the sensing surface. Thus,to minimize this dampening, it is preferable to expose only that part ofthe resonator required to perform the measurement to the liquid testsample, which again is the sensing electrode surface.

The earliest and most commonly used method for ensuring only oneelectrode of an oscillating crystal resonator came into contact with theliquid test sample was to sandwich, or “mount”, the resonator between apair of rubber O-rings or gaskets. The O-ring or gasket on the sensingsurface of the resonator was then interfaced with a well or cell suchthat the sample solution can be applied to that surface without beingexposed to the other parts of the resonator. An example of this doubleO-ring resonator mounting configuration is disclosed in U.S. Pat. No.5,135,852. As noted by the patent's author, a problem of this resonatormounting configuration is that the O-rings or gaskets exert fluctuatingand non-reproducible pressure on the oscillating resonator, whichdirectly impacts the sensitivity of the resonator. More specifically theauthor stated: in this sensor structure, the seals are placed at theedge of the sensor where the interference with its oscillations isminimal. However, this setup has the following drawbacks: 1) sensorresponse is strongly influenced by mounting pressure of the sample fluidwithin the cell, and this pressure adjustment is not readilyreproducible; 2) during assembly, the quartz plate is handled directlyresulting in the risk of damaging the fragile quartz plate; and 3) evenwhen fixed firmly between the O-rings, distortions due to pressurefluctuations in the tested liquid, and expansion or contractions of theplate due to thermal changes will stress the sensor plate and causefriction between the sensor and the O-ring which in turn will result indecreased Q-factor and unsteady oscillations, i.e. noise sensorresponse.

This description highlights one of the basic problems in the use ofoscillating crystal resonators in liquid-based applications. To beeffectively employed as a liquid phase biosensor, the oscillatingcrystal resonator needs to be physically interfaced with liquid deliveryunit so that only the sensing surface comes in contact with the liquidsample. However, any and all physical contact with the resonator, i.e.,from the sample fluid, the mounting structure, e.g., the O-rings, etc.,dampens its oscillation freedom and thus lowers it sensitivity andoverall functionality. Similar to the example discussed above, the vastmajority of resonator mounting designs to date have employed the use ofelastic mounting seals or elastic adhesives to hold the resonator inplace and create the liquid tight seal for the test sample chamber. Theargument for the continued use of flexible mounting materials is thebelief that the elasticity of the seal or adhesive will minimize itsdampening of the resonator oscillation. However, while this elasticityin the mounting material may minimize dampening, it clearly has animpact on reproducibility between individual measurements on a singleresonator and for measurements between different resonators.

To reduce the amount of stress on the mounted resonator, reduce thesignal noise, and to improve reproducibility, various assemblies havesubsequently been developed which use only one flexible O-ring or gasketand secure the resonator by pressing it against a solid mount, such asfor example those sensor structures disclosed in PCT Application Nos.WO/2004/040268, WO/2002/061396, and WO/2002/012873. In all of thesedesigns, the sensing (front) surface of the resonator is in physicalcontact with parts, solid or flexible, of the mounting assembly.However, because the resonator oscillations propagate out to the edge ofthe piezoelectric plate (even if the coated electrode does not reach tothe edge of the plate), any component making physical contact with thesensing surface will impact the resonator response. Also, even slightdistortions of the resonator from mechanical or thermal variations ofthe mounting assembly will result in noise added to the sensor response.As a result, none of the designs disclosed in the applicationsfacilitates the construction of a reproducible sensor assembly.

More recently, another form of resonator mount designed to reduce sensornoise was disclosed in PCT Application No. WO/2002/047246. In contrastto the prior art sensor designs, in this design the resonator is placedwith its non-sensing surface on a solid support surface and fixed to thesurface with a flexible adhesive applied only along the edge of theresonator plate. Because the resonator is not exposed to direct andvariable physical pressure from clamping, and the sensing electrode isnot in contact with any mounting components, a significant noisereduction for liquid sensing applications is achieved, while at the sametime the sensing electrode is isolated from the driving electrode,thereby preventing a short circuit across the resonator plate. However,this improved design still suffers from a number of problems that havebeen discussed in literature publications released following the patentfiling. In particular, in practice it is very difficult to uniformlyapply flexible adhesive around the edge of the very thin resonatorplates (˜100 um) without depositing some amount of adhesive onto thesensing surface. It is also known that the resonator oscillations willreflect from the secured edge of the resonator plate, and thus theadhesive placed around the edge of the plate will still have an impacton the resonator response. Furthermore, while it is not physicallypressed against the mounting substrate, the entire non-sensing surfaceof the resonator is in physical contact with the mounting substrate.Thus, even slight imperfections on the mounting surface will result inirregular stress on the resonator and noise in the sensor response.

Another problem with the use of oscillating crystal resonators in liquidbased applications is the signal noise and response reproducibilityissues arising from changes in hydraulic pressure on the resonatorsurface(s) during sample analysis. A primary requirement for the use ofmounted oscillating crystal resonators in liquid based applications isthe ability to expose sample-containing liquids to the sensing surfaceof the resonator as discrete volumes or plugs within a continuouslyflowing stream of a sample-less liquid. In a wide variety ofapplications, it is highly beneficial for the resonator sensing surfaceto be exposed to a liquid solution identical to that of the sample butwithout the sample before and after the sample solution is applied tothe resonator sensing surface. It is also highly advantageous in a widevariety of applications that the exchanges from non-sample solution, tosample containing solution, and back to non-sample solution happen asinstantaneously as possible. The processes involved with executing theadditions and exchanges of sample and non-sample solutions to thesensing surface of the resonator result in pressure changes on theresonator surface(s), often resulting in non-reproducible stress on theresonators and dampening of the crystal oscillations. These stresses anddampening events can greatly diminish the sensitivity andreproducibility of the resonators as mass measurement sensors.

One approach to dealing with this problem has been to apply acounter-acting pressure on the backside of the resonator (drivingelectrode surface) to minimize irregular stress, deviations, andfluctuations on the sensing surface of the resonator. A problem withthis type of solution comes from the design constraints of applyingpressure on the backside of the mounted resonator due to the presence ofthe electrodes and the detection electronics connected to thoseelectrodes. Also the intrinsic dampening of the resonator oscillationsfrom exerting pressure on both faces of the resonator, and thedifficulty in accurately matching the counter pressure on the drivingelectrode surface to the pressure being exerted on the sensing surfaceby the sample fluid or solution, make this practice highly impractical.

Thus, while the current resonator mounting designs are functional, theyare far from optimal. Often the influence from erroneous signals in theform of response dampening, random noise, and drift created by theresonator mounting process lowers the sensitivity and reproducibility ofthe resonator to such a degree that the technology is not feasible forthe majority of desired applications as a liquid based sensor. Clearly,to advance the use of this technology there is a requirement for animproved resonator mounting design. Ideally the mounting design wouldaffix the oscillating crystal resonator to the mounting substrate in ahighly stable, highly reproducible manner, fully isolating the liquidsample exposure to only the sensing surface of the resonator, whilerequiring only a minimal amount of physical contact between the mountingand sample delivery assembly and the oscillating resonator surfaces.

SUMMARY OF THE INVENTION

According to one aspect of the present invention, a sensor is providedwith a crystal resonator that is affixed to a substrate for the sensorwith an adhesive that forms a rigid bond between the resonator and thesubstrate when cured. The rigid bond between the resonator and thesubstrate effectively causes the resonator to vibrate in a manner thatis not affected by the connection of the resonator to the substrate.This rigid connection greatly enhances the reproducibility of thevibration that can be generated within the resonator by effectivelyisolating the resonator from the substrate. In other words, the rigidconnection between the resonator and the substrate prevents anydampening of the vibrations within the resonator as a result of thisconnection, as opposed to prior art flexible connections between thesubstrate and resonator. Further, because the resonator is rigidlyaffixed to the substrate, during operation the resonator will not“creep” or shift position with regard to the substrate. As such, theresonator will remain in the same position throughout multiple uses ofthe sensor, such that the vibration characteristics of the resonatorwill not change, further adding to the reproducibility of the vibrationsin the resonator.

According to another aspect of the present invention, the adhesiveforming the rigid bond between the resonator and the substrate ispresent around the entire periphery of the resonator. This ensures thatthe vibration characteristics of the resonator are uniform across theentire resonator, to prevent any variations in these vibrationcharacteristics of the resonator.

According to still another aspect of the invention, the resonator ismounted to a substrate of a sensor that enables the sample fluid to beinjected into a stream of a control fluid already passing over theresonator. To offset the increased pressure exerted on the overall fluidstream passing over the resonator from the introduction of the samplefluid and avoid any inaccurate affects on the vibration of theresonator, the streams of the control fluid can be reduced in volume.This maintains the overall pressure of the fluid streams passing overthe resonator at a constant level, such that the only effects on thevibration of the resonator are produced by the sample fluid.

Numerous other aspects, features and advantages of the present inventionwill be made apparent from the following detailed description takentogether with the drawing figures.

BRIEF DESCRIPTION OF THE DRAWINGS

The drawing figures illustrate the best mode currently contemplated ofpracticing the present invention.

In the drawings:

FIG. 1 is a cross-sectional view of a sensor mounting assemblyconstructed according to the present invention;

FIG. 2 is a top plan view of the assembly of FIG. 1;

FIG. 3 is a cross-sectional view of a second embodiment of the sensormounting assembly of the present invention; and

FIG. 4 is a top plan view of a number of laminar fluid streams flowingover a sensor mounting assembly constructed according to the presentinvention.

DETAILED DESCRIPTION OF THE INVENTION

With reference now to the drawing figures in which like referencenumerals represent like parts throughout the disclosure, a sensorassembly constructed according to the present invention is illustratedgenerally at 100 in FIG. 1. In one embodiment for the assembly shown inFIG. 1, the assembly 100 is a dry side sensor mounting assembly for asingle sensing spot crystal resonator 30 assembled into a sampledelivery flow cell 1000. The assembly 100 is formed with a substrate 10of a fluid-impervious and non-conductive material that includes acentral bore 20 and a pair of electrode contact bores 22, 24 disposed onopposite sides of the central bore 20, though the bores 22, 24 can belocated in any suitable location around the bore 20. Additionally, thecentral bore 20 and the bores 22, 24 are shaped as desired toaccommodate the particular structure of the assembly 100, and thereforecan have any desired shape.

An oscillating crystal resonator 30 formed of a conventional quartzcrystal material is positioned on the upper surface 62 of the substrate10, and includes a driving electrode 32 and a sensing electrode 34disposed on opposite sides of the resonator 30. The driving electrode 32is positioned adjacent the upper surface 62 of the substrate 10, whilethe sensing electrode 34 is primarily positioned on the surface of theresonator 30 opposite the driving electrode 32. However, the sensingelectrode 34 does include a wrap-around portion 36 that extends from thesensing electrode 34 around the resonator 30 onto the surface of theresonator 30 against which the driving electrode 32 is positioned.However, the wrap-around portion 36 of the sensing electrode 34terminates at a point spaced from the driving electrode 32, and does notcontact the driving electrode 32 to prevent a short circuit across theelectrodes 32 and 34 on the resonator 30.

The bores 22, 24 are each filled with a conductive adhesive plug 50 thatextend a short distance above the upper surface 62 of the substrate 10.The portion of these plugs 50 that extends above the upper surface 62 ofthe substrate 10 contacts the driving electrode 32 and the wrap-aroundportion 36 of the sensing electrode 34, respectively. Opposite theelectrodes 32 and 34, the plugs 50 are each operably connected with oneof a pair of conductive tracks 60 formed on the lower surface 64 of thesubstrate 10. The conductive tracks 60 formed on the lower surface 64 ofthe substrate 10 can be formed in any conventional manner, such as bybeing printed on the substrate 10 with a conductive ink, and terminateat electrical contacting pads 68 at the edges of the lower surface 64 ofthe substrate 10, enabling the mounted resonator 30 and the electrodes32, 34 thereon to be easily interfaced with corresponding detectionelectronics systems, as are known in the art.

When positioned on the substrate 10, the driving or non-sensingelectrode 32 of the resonator 30 faces the mounting substrate 10, and ispositioned directly over the bore 20 such that the majority of thedriving electrode 32 is not physically contacting the mounting substrate10. The resonator 30 is also positioned onto the mounting substrate 10such that only the terminal end of the end wrap-around portion 36 of theelectrode 34 is in contact with the substrate 10. For each electrode 32and 34, the only actual part of the electrodes 32 and 34 that is indirect contact with the substrate 10 are those portions in contact withthe conductive adhesive plugs 50 and a rigid adhesive 40, to bedescribed. For other embodiments for mounting resonators 30 possessingmultiple sensing spots, and thus multiple driving electrodes 32 and/orsensing electrodes 34, the mounting substrate 10 can include multiplebores 20 for the driving electrodes 32, and multiple bores 22 and 24 foraccommodating a number of conductive adhesive plugs 50, with one bore22, 24 and corresponding plug 50 for each driving electrode 32 andsensing electrode 34. Furthermore, the bores 20-24 will be of a sizesuch that the majority of each of the driving electrodes 32 and sensingelectrodes 34 do not physically contact the substrate 10 when theresonator 30 is mounted thereto.

The resonator 30 with electrodes 32 and 34 is mounted, or affixed, ontothe substrate 10 such that the electrodes 32 and 34 are disposed incontact with the plugs 50 using a very low viscosity, non-conductive,rigid bond-forming, and solvent resistant adhesive 40. One example of anadhesive that is suitable for use as the adhesive 40 is the adhesivesold under the trade name Loctite 3491, by Henkel of Düisseldorf,Germany. In one method of mounting the resonator 30 to the substrate 10,a small amount of adhesive 40 is deposited onto the upper surface 62immediately around the driving electrode bores 20, 22, and 24 in thesubstrate 10. The resonator 30 is placed onto the substrate 10 with thedriving electrode 32 facing the substrate 10 and in such a manner thatthe adhesive 40 remains only in the areas between the resonator 30 andthe upper surface 62 of the substrate 10. The low viscosity of theadhesive 40 facilitates capillary distribution of the adhesive 40 toform a uniform thin film over the entire area of the upper surface 62 ofthe substrate 10 between the substrate 10 and resonator 30. The verysmall volume of adhesive 40, in part due to the particular properties ofthe adhesive 40, will not spread across the resonator 30 in areas wherethe substrate 10 has bores 20, 22, and 24, due to lack of capillaryaction. Therefore, the adhesive 40 does not cover any of the adjacentsurface area of the driving electrode 32 or the wrap-around portion 36of the sensing electrode 34 of the resonator 30 disposed across thecentral bore 20, or the electrode bores 22, 24 in the substrate 10. Thisdistribution of the adhesive 40 between the resonator 30 and themounting substrate 10 forms a rigid and liquid-tight seal around thecentral bore 20 in the substrate 10, and around all of the electrodebores 22, 24 also formed in the substrate 10, which are disposedpreferably immediately adjacent the central bore 20. When cured in asuitable manner, such as by heat, UV light or over time, the adhesive 40creates a rigid permanent seal mounting the resonator 30 to thesubstrate 10 while minimizing any physical contact of the adhesive 40 orthe substrate 10 with the driving electrode 32 and sensing electrode 34on the resonator 30. With this construction, the resonator 30 is rigidlysecured to the substrate 10 in a manner that prevents the fluid beingsensed by the sensing electrode 34 from also coming into contact withthe driving electrode 32 and creating a short between the electrodes 32and 34. Further, because the resonator 30 is mounted to the substrate 10in rigid manner that minimizes the contact of the resonator 30 and theelectrodes 32 and 34 with the substrate 10, the assembly 100simultaneously isolates, to the extent possible, the electrodes 32 and34 from the substrate 10, and provides a stable attachment of theresonator 30 to the substrate 10 to maintain the vibration properties ofthe resonator 30 constant.

When mounted in this manner using the adhesive 40, the electrodes 32 and34 of the resonator 30 are also connected to the conductive tracks 60 onthe lower surface 64 of the substrate 10 by the plugs of conductiveadhesive 50 subsequently formed in the electrode bores 22, 24 present inthe substrate 10. The conductive adhesive, such as the adhesive soldunder the trade name Silver Conductive Paint by RS Components Ltd ofAuckland, NZ, is applied into the bores 22 and 24 after attachment ofthe resonator 30 m to the substrate 10 to form the plugs 50 in a mannersuch that that, when fully formed, each plug 50 contacts one of theresonator electrodes 32 and 34 at one end through the open areas in themounting adhesive 40 remaining above the bores 22, 24 in the substrate10, and the conductive tracks 60 on the formed on the lower surface 64of the substrate 10 opposite the electrodes 32, 34 at the opposite end.

In order to form the assembly 100 into a flow cell 1000 for delivery ofsample fluids to the resonator 30 for analysis, the assembly 100 has aliquid sealing gasket 70 of a known height positioned on the substrate10 around the resonator 30. The gasket 70 can be formed of any desiredmaterial, with a fluid-impervious solvent resistant rubber or siliconebeing especially preferred. Subsequently, a fluid delivery block 80formed of a material similar to that used for the substrate 10 issecured to the gasket 70 opposite the substrate 10. The fluid deliveryblock 80 is affixed to the gasket 70 in any suitable fluid-tight mannerto enclose and define the interior 85 of the flow cell 1000, and isshaped to have a number of fluid delivery ports 90 formed therein. Theshape and height of the sealing gasket 70 determines the shape and totalvolume of the flow cell 1000, and is selected such that neither thesealing gasket 70 nor the fluid delivery block 80 will make physicalcontact with the mounted resonator 30 when the gasket 70 and fluiddelivery block 80 are engaged with the substrate 10, such as by clampingthe gasket 70 and block 80 to the substrate 10. In addition, thepositions of the fluid delivery ports 90 on the fluid delivery block 80are disposed relative to the shape of the sealing gasket 70 such thatfluid samples entering the cell must pass over the sensing electrode 34on the mounted resonator 30 prior to exiting the cell 1000. The numberof ports 90 formed in the block 80 can be selected as desired based onthe number of fluid sources that are to be introduced into the flow cell1000. Also, the positions of the ports 90 can be located such that thefluid streams introduced into the flow cell can be removed from the cell1000 after passing over the resonator 30.

Referring now to FIG. 2, in an alternative embodiment of theconstruction of the driving electrode 32 on the resonator 30, thedriving electrode 32 is designed so that only the ‘neck’ of the drivingelectrode 32 on the resonator 30 physically contacts the mountingsubstrate 10. In this configuration, the amount of the surface of thedriving electrode 32 that contacts the substrate 10 via the adhesive 40and the plug 50 is further minimized

Looking now at FIG. 3, another alternative construction for the assembly100′ is illustrated that utilizes an adhesive channel 140. This assembly100 is very similar to the assembly 100′ illustrated in FIG. 1, with thekey difference being the fact that the mounting adhesive 40 that bondsthe resonator 30 to the mounting substrate 10 is contained within anetched channel 140 cut into the upper surface 62 of the of the substrate10. By confining the mounting adhesive 40 to the etched channel 140 whenthe resonator 30 is mounted, the portion of the driving electrode 32 onthe resonator 30 that is to be connected to the plug 50 partially restsflush against the upper surface 62 of the mounting substrate 10. The useof the adhesive channel 140 ensures that this portion of the drivingelectrode 32 contacts the upper surface 62 of the mounting substrate 10in a uniform and rigid manner, which results in more uniform andreproducible pressure on the driving electrode 32. The channel 140 alsohelps to ensure the entire surface area of sensing electrode surface 34is an equidistant height from the upper surface 62 of the mountingsubstrate 10. Thus, when the mounted resonator 30 is interfaced with afluid delivery block 80 and gasket 70 to form a flow cell 1000, thespace between the fluid delivery block 80 and the entire exposed surfaceof the sensing electrode 34 is uniform.

To mount the resonator 30 onto a mounting substrate 10 possessing anadhesive channel 140, an amount of the adhesive 40 sufficient to contactthose parts of the resonator 30 that are to be located directly over thechannel 140 is deposited into the channel 140. The resonator 30, withdriving electrode 32 facing the mounting substrate 10, is placed ontothe substrate 10 so that the majority of the driving electrode 32 lieswithin or over the central bore 20 which is circumscribed by the channel140, and the driving and sensing electrodes 32 and 34 are located inalignment with the conductive adhesive bores 22 and 24 and alsocircumscribed by the channel 140. Any excess adhesive 40 present in thechannel 140 during the mounting process will escape through a number ofadhesive exhaust holes 150 formed in the substrate 10 below the channel140. Alternatively, the resonator 30 can first be properly positionedonto the mounting substrate 10 over and/or partially within the channel140, and then mounting adhesive 40 can be injected into the adhesivechannel 140 through one or more of the adhesive exhaust holes 150 withany excess adhesive 40 escaping through the other exhaust holes 150. Asin previous embodiments, when the adhesive 40 cures, it contracts,slightly pulling the resonator 30 into tight engagement against themounting substrate 10 and forms a rigid, liquid tight bond between theresonator 30 and substrate 10. However, the width of the adhesivechannel 140 in relation to the size of the resonator 30 is in a ratiothat is small enough not to cause the resonator 30 to bend as the curingadhesive 40 contracts. After the adhesive 40 has sufficiently cured, theconductive adhesive plugs 50 can then be formed within the bores 22 and24 to connect the electrodes 32 and 34 on the resonator 30 to theconductive tracks 60 on the lower surface 64 of the substrate 10.

After the assembly 100 and the flow cell 1000 have been constructed inthe above manner, it is then possible to pass a fluid stream through theflow cell 1000 in order to allow variations in the vibrations of theresonator 30 caused by materials in the fluid attaching to the resonator30 to be detected, and thus obtain information concerning the componentsof the fluid sample flowing over the resonator 30. To do so, it isnecessary to pass the fluid sample to be analyzed directly over theresonator 30 when it is injected into the flow cell 1000. It is knownthat when two or more independent streams of fluid flowing underconditions of laminar flow, i.e., a low Reynolds number for the flow,are in direct contact with each other and flow in the same direction,i.e. flow parallel to one another, there will be no mixing of the fluidsother than by diffusion. Also, by varying the rates of flow of thedifferent fluid streams in relation to each other, the size and positionof the streams can be altered, as disclosed in Biosensors andBioelectronics Vol. 13 No. 3-4, pages 47-438, 1998, such that the fluidstreams can be moved within the space through which they are flowing bychanging the relative flow rates. It is also known that oscillatingcrystal resonators, such as quartz crystal resonators, possess an areaof higher detection sensitivity based on the dimensions of the drivingelectrode of the resonator in relation to the sensing electrode, asdiscussed in Ward and Delawski, Anal. Chem. 1991, 63, 886-890. Thus,focusing the flow of a sample fluid stream over this detection ‘sweetspot’ provides a benefit in the use of oscillating crystal resonators 30as sensors by minimizing the amount of sample consumption needed toachieve the most efficient level of detection sensitivity.

Referring now to FIG. 4, a close up section of the previous describeflow cell embodiments where only a flow cell area 1000 is illustratedthat includes the sensing electrode 34 of a substrate mounted resonator30 in any one of the above-described manners, and a fluid delivery block80 disposed above the resonator 30 and engaged with the substrate by agasket 70. In FIG. 4, the fluid delivery block 80 is not shown as in theprevious embodiments, but it is implied that the various fluid streams300, 400 and 500 are introduced into the flow cell 1000 through therespective fluid delivery ports 90 located in the delivery block 80. InFIG. 4 the substrate onto which the resonator 30 is mounted is not shownin detail as in previous embodiment, but is implied to constitute theentire surface area beneath the resonator 30 and within the gasket 70boundaries. The two guide fluid streams 300 and 400 are continuouslydirected into the flow cell 1000 prior to the introduction of any samplefluid stream 500. These guide fluid streams 300 and 400 also provide abaseline hydrostatic pressure exerted onto the sensing electrode 34 ofthe resonator 10. When the sample fluid stream 500 is introduced intothe flow cell 1000 through one of the fluid delivery ports 90, the guidefluid streams 300 and 400 are able to control the position and width ofthe sample or reagent fluid stream 500 through the flow cell 1000 viathe process of hydrodynamic focusing described previously. Bycontrolling the flow rate of the guide fluid streams 300 and 400relative to that of the sample or reagent fluid stream 500, it ispossible to direct or position the sample or reagent fluid stream 500over the portion of the sensing electrode 3 of the resonator 30 that ispositioned directly over the driving electrode 32, located on the backside of the resonator 30) or the ‘sweet spot’ of detection defined bythe relative positions of the electrodes 32 and 34 located on eitherface of the resonator 30.

In addition, as with any enclosed chamber, the addition or removal offluid from that chamber will change the hydrostatic pressure on allwalls of the chamber. In the above described assembly where the flowcell 1000 is created over a substrate to which the resonator 30 ismounted in any of the previously described manners, the addition orremoval of one or more of the fluid streams 300, 400 or 500 from theenclosed flow cell 1000 will change the hydrostatic pressure on thesurface of the resonator 30, and thus alter its detection response.Therefore, to maintain a constant level of hydrostatic pressure on themounted resonator 30 within an enclosed flow cell 1000 during thedelivery or removal of sample or reagent fluid streams 300, 400 and/or500 to the sensing surface 34 of the resonator 30, when the flow cell1000 constructed according to the present invention is operated toinject a sample fluid stream 500 into the cell 1000, the flow rates ofthe constantly flowing guide fluid streams 300 and 400 are altered tocompensate for this pressure change. More particularly, during normaloperation of the flow cell 1000, the guide fluid streams 300 and 400 areintroduced through one or more of the fluid delivery ports 90 tocontinuously flow through the flow cell 1000 defined by the mountingsubstrate, the fluid delivery block 80 and the liquid sealing gasket 70.These guide fluid streams 300 and 400 create a specific and constantpressure upon the sensing electrode 34 of the mounted resonator 30within the flow cell 1000 based on their rate of flow. When introducinga sample fluid stream 500 into the flow cell 1000 through a separatefluid delivery port 90 for analysis, the flow rate of the guide fluidstreams 300 and 400 already flowing in the cell 1000 are lowered by arate corresponding or equivalent to the rate of flow of the newlyintroduced fluid stream 500. This adjustment is in addition to theadjustment of the flow rates of the guide fluid streams 300 and 400relative to the flow of the sample fluid stream 500 to direct the samplefluid stream 500 over the sensing electrode 34 in such a manor that itis focused on a path equivalent to the width of the driving electrode 32on the backside of the resonator 10. When the flow of the sample fluidstream 500 is stopped, the flow rates of the guide fluid streams 300 and400 are increased to compensate for the loss of fluid entering the cell1000, and maintain the constant pressure level on the sensing electrode3 of the resonator 30. In this manner, when all of the various fluidstreams 300, 400 and 500 are generally equal in their composition ordensity of the carrier fluid from which they are formed, the hydrostaticstatic pressure within the cell 1000 is held constant to achieve aconsistent response from the resonator 30 when analyzing a sample fluid.

Various alternatives are contemplated as being within the scope of thefollowing claims particularly pointing out and distinctly claiming thesubject matter regarded as the invention.

1. A sensor comprising: a) a substrate including at least one boreextending therethrough; and b) an oscillating sensing device rigidlymounted to one surface of the substrate over the bore, the sensingdevice including a first electrode disposed in communication with the atleast one bore and a second electrode disposed on the oscillatingsensing device opposite the first electrode, wherein the oscillatingsensing device is affixed to the substrate by an adhesive positionedaround the periphery of the at least one bore, and wherein the adhesiveis disposed within a channel formed in the substrate around theperiphery of the at least one bore.
 2. The sensor of claim 1 furthercomprising at least one adhesive exit port extending through thesubstrate into connection with the channel.
 3. A sensor comprising: a) asubstrate including at least one bore extending therethrough; and b) anoscillating sensing device rigidly mounted to one surface of thesubstrate over the bore, the sensing device including a first electrodedisposed in communication with the at least one bore and a secondelectrode disposed on the oscillating device opposite the firstelectrode, wherein each of the first and second electrodes are operablyconnected to a conductor passing through the substrate and the adhesiveand wherein the first electrode is disposed positioned directly over andcircumscribed by the at least one bore except at the point wherecontacted by the conductor.
 4. A flow cell for use in analyzing fluidsamples, the flow cell comprising: a) sensor comprising a substrateincluding at least one bore extending therethrough and an oscillatingsensing device rigidly mounted to one surface of the substrate over thebore, the sensing device including a first electrode disposed incommunication with the at least one bore and a second electrode disposedon the oscillating sensing device opposite the first electrode; b) asealing member secured in a fluid-tight manner to the substrate aroundthe oscillating sensing device; and c) a flow deliver block secured tothe sealing member in a fluid-tight manner opposite the substrate andincluding a number of fluid dispensing ports formed therein, wherein theoscillating sensing device is mounted by an adhesive flush against thesurface of the substrate to completely cover the at least one bore.
 5. Aflow cell for use in analyzing fluid samples, the flow cell comprising:a) sensor comprising a substrate including at least one bore extendingtherethrough and an oscillating sensing device rigidly mounted to onesurface of the substrate over the bore, the sensing device including afirst electrode disposed in communication with the at least one bore anda second electrode disposed on the oscillating sensing device oppositethe first electrode; b) a sealing member secured in a fluid-tight mannerto the substrate around the oscillating sensing device; and c) a flowdeliver block secured to the sealing member in a fluid-tight manneropposite the substrate and including a number of fluid dispensing portsformed therein, wherein the sealing member and the flow delivery blockdo not contact the oscillating sensing device.
 6. A method of assemblinga sensor comprising the steps of: a) providing an oscillating sensingdevice having a first electrode disposed on one side of the device and asecond electrode disposed on the device opposite the first electrode;and b) rigidly securing the oscillating device to a surface of asubstrate such that the device completely covers at least one boreformed through the substrate and such that the first electrode ispositioned substantially within the at least one bore, wherein the stepof rigidly securing the oscillating device to the substrate comprisesthe steps of: i) applying an adhesive to one of a periphery of theoscillating sensing device or a periphery of the at least one bore inthe substrate; and ii) pressing the oscillating sensing device and thesubstrate into engagement with one another.
 7. The method of claim 6further comprising the step of forming a conductor that is operablyconnected to one of the first electrode or the second electrode throughthe substrate after pressing the oscillating sensing device and thesubstrate into engagement with one another.
 8. The method of claim 6further comprising the steps of: a) securing a sealing member to a fluiddelivery block to define an interior within the sealing membersufficient to completely encompass the oscillating sensing mechanism;and b) engaging the sealing member with the substrate around theoscillating sensing mechanism.
 9. A method for analyzing a fluid sampleutilizing a flow cell including a substrate including at least one boreextending therethrough, and an oscillating sensing device rigidlymounted to one surface of the substrate over the bore, the sensingdevice including a first electrode disposed in communication with the atleast one bore and a second electrode disposed on the oscillatingsensing device opposite the first electrode and a fluid delivery blockaffixed to and spaced from the substrate over the oscillating sensingdevice by a sealing member, the method comprising the steps of: a)introducing at least one guide fluid stream into the flow cell throughthe fluid delivery block to flow over the oscillating sensing device togenerate a hydrostatic pressure value within the flow cell; b)introducing at least one sample fluid stream into the flow cell throughthe flow delivery block at a location separate from the at least oneguide fluid stream to flow over the oscillating sensing device; and c)reducing the flow rate of the at least one guide fluid streamsimultaneously with the introduction of the at least one sample fluidstream to maintain the hydrostatic pressure value within the flow cellconstant.
 10. The method of claim 9 wherein the step of introducing theat least one guide fluid stream comprises introducing at least two guidefluid streams into the flow cell through spaced locations in the fluiddelivery block; and wherein the step of reducing the flow rate of the atleast one guide fluid stream comprises varying the flow rate of at leastone of the at least two guide fluid streams to shift a flow path of theat least one sample fluid stream within the flow cell simultaneouslywith maintaining constant the hydrostatic pressure value within the flowcell.
 11. A method for analyzing a fluid sample utilizing a flow cellincluding a sensing device disposed within the flow cell and a fluiddelivery block configured to direct multiple fluid streams into the flowcell over the sensing device, the method comprising the steps of: a)introducing at least one guide fluid stream into the flow cell throughthe fluid delivery block to flow over the sensing device to generate ahydrostatic pressure value within the flow cell; b) introducing a samplefluid stream into the flow cell through the flow delivery block at alocation separate from the at least one guide fluid stream to flow overthe sensing device; and c) reducing the flow rate of the at least oneguide fluid stream simultaneously with the introduction of the samplefluid stream to maintain the hydrostatic pressure value within the flowcell constant.